Patent application title: SOYBEAN VARIETY A1016008

Abstract:

The invention relates to the soybean variety designated A1016008. Provided
by the invention are the seeds, plants and derivatives of the soybean
variety A1016008. Also provided by the invention are tissue cultures of
the soybean variety A1016008 and the plants regenerated therefrom. Still
further provided by the invention are methods for producing soybean
plants by crossing the soybean variety A1016008 with itself or another
soybean variety and plants produced by such methods.

Claims:

1. A seed of soybean variety A1016008, representative seed of said soybean
variety having been deposited under ATCC Accession No. ______.

2. A plant of soybean variety A1016008, representative seed of said
soybean variety having been deposited under ATCC Accession No. ______.

7. A soybean plant regenerated from the tissue culture of claim 5, wherein
the regenerated soybean plant expresses all of the physiological and
morphological characteristics of the soybean variety A1016008,
representative seed of said soybean variety having been deposited under
ATCC Accession No. ______.

8. A method of producing soybean seed, comprising crossing a plant of
soybean variety A1016008 with itself or a second soybean plant,
representative seed of said soybean variety having been deposited under
ATCC Accession No. ______.

9. The method of claim 8, further defined as a method of preparing hybrid
soybean seed, comprising crossing a plant of soybean variety A1016008
with a second, distinct soybean plant, representative seed of said
soybean variety having been deposited under ATCC Accession No. ______.

10. An F1 hybrid seed produced by the method of claim 9.

11. A method of producing a plant of soybean variety A1016008 comprising
an added desired trait, the method comprising introducing a transgene
conferring the desired trait into a plant of soybean variety A1016008,
representative seed of said soybean variety having been deposited under
ATCC Accession No. ______.

13. The method of claim 12, wherein the desired trait is herbicide
tolerance and the tolerance is conferred to an herbicide selected from
the group consisting of glyphosate, sulfonylurea, imidazalinone, dicamba,
glufosinate, phosphinothricin, phenoxy proprionic acid, cyclohexone,
triazine, benzonitrile and broxynil.

14. The method of claim 11, wherein the desired trait is insect resistance
and the transgene encodes a Bacillus thuringiensis (Bt) endotoxin.

17. A method of introducing a single locus conversion into soybean variety
A1016008 comprising:(a) crossing a plant of soybean variety A1016008 with
a second plant comprising a desired single locus to produce F1 progeny
plants, representative seed of said soybean variety having been deposited
under ATCC Accession No. ______.(b) selecting at least a first progeny
plant from step (a) that comprises the single locus to produce a selected
progeny plant;(c) crossing the selected progeny plant from step (b) with
a plant of soybean variety A1016008 to produce at least a first backcross
progeny plant that comprises the single locus; and(d) repeating steps (b)
and (c) with the selected backcross progeny plant from step (d) used in
place of the first progeny plant of step (b) during said repeating,
wherein steps (b) and (c) are repeated until at least a first backcross
progeny plant is produced comprising the single locus and essentially all
of the physiological and morphological characteristics of soybean variety
A1016008 when grown in the same environmental conditions.

19. The method of claim 18, wherein the trait is tolerance to an herbicide
selected from the group consisting of glyphosate, sulfonylurea,
imidazalinone, dicamba, glufosinate, phenoxy proprionic acid,
cyclohexone, triazine, benzonitrile and broxynil.

20. The method of claim 18, wherein the trait is insect resistance and the
insect resistance is conferred by a transgene encoding a Bacillus
thuringiensis endotoxin.

22. A plant of soybean variety A1016008, further comprising a single locus
conversion, wherein the single locus conversion is introduced into
soybean variety A1016008 by backcrossing or genetic transformation,
representative seed of said soybean variety having been deposited under
ATCC Accession No. ______.

23. A method of producing an inbred soybean plant derived from the soybean
variety A1016008, the method comprising the steps of:(a) preparing a
progeny plant derived from soybean variety A1016008 by crossing a plant
of the soybean variety A1016008 with a soybean plant of a second variety,
representative seed of said soybean variety having been deposited under
ATCC Accession No. ______;(b) crossing the progeny plant with itself or a
second plant to produce a seed of a progeny plant of a subsequent
generation;(c) growing a progeny plant of a subsequent generation from
said seed and crossing the progeny plant of a subsequent generation with
itself or a second plant; and(d) repeating steps (b) and (c) until an
inbred soybean plant derived from the soybean variety A1016008 is
produced.

24. A method of producing a commodity plant product comprising obtaining
the plant of claim 2 or a part thereof and producing said commodity plant
product therefrom.

Description:

[0001]This application claims the priority of U.S. Provisional Appl. Ser.
No. 61/226,274, filed Jul. 16, 2009, the entire disclosure of which is
incorporated herein by reference.

BACKGROUND OF THE INVENTION

[0002]1. Field of the Invention

[0003]The present invention relates generally to the field of soybean
breeding. In particular, the invention relates to the novel soybean
variety A1016008.

[0004]2. Description of Related Art

[0005]There are numerous steps in the development of any novel, desirable
plant germplasm. Plant breeding begins with the analysis and definition
of problems and weaknesses of the current germplasm, the establishment of
program goals, and the definition of specific breeding objectives. The
next step is selection of germplasm that possess the traits to meet the
program goals. The goal is to combine in a single variety an improved
combination of desirable traits from the parental germplasm. These
important traits may include higher seed yield, resistance to diseases
and insects, better stems and roots, tolerance to drought and heat,
better agronomic quality, resistance to herbicides, and improvements in
compositional traits.

[0006]Soybean, Glycine max (L.), is a valuable field crop. Thus, a
continuing goal of plant breeders is to develop stable, high yielding
soybean varieties that are agronomically sound. The reasons for this goal
are to maximize the amount of grain produced on the land used and to
supply food for both animals and humans. To accomplish this goal, the
soybean breeder must select and develop soybean plants that have the
traits that result in superior varieties.

SUMMARY OF THE INVENTION

[0007]One aspect of the present invention relates to seed of the soybean
variety A1016008. The invention also relates to plants produced by
growing the seed of the soybean variety A1016008, as well as the
derivatives of such plants. Further provided are plant parts, including
cells, plant protoplasts, plant cells of a tissue culture from which
soybean plants can be regenerated, plant calli, plant clumps, and plant
cells that are intact in plants or parts of plants, such as pollen,
flowers, seeds, pods, leaves, stems, and the like.

[0008]Another aspect of the invention relates to a tissue culture of
regenerable cells of the soybean variety A1016008, as well as plants
regenerated therefrom, wherein the regenerated soybean plant is capable
of expressing all the physiological and morphological characteristics of
a plant grown from the soybean seed designated A1016008.

[0009]Yet another aspect of the current invention is a soybean plant
comprising a single locus conversion of the soybean variety A1016008,
wherein the soybean plant is otherwise capable of expressing all the
physiological and morphological characteristics of the soybean variety
A1016008. In particular embodiments of the invention, the single locus
conversion may comprise a transgenic gene which has been introduced by
genetic transformation into the soybean variety A1016008 or a progenitor
thereof. In still other embodiments of the invention, the single locus
conversion may comprise a dominant or recessive allele. The locus
conversion may confer potentially any trait upon the single locus
converted plant, including herbicide resistance, insect resistance,
resistance to bacterial, fungal, or viral disease, male fertility or
sterility, and improved nutritional quality.

[0010]Still yet another aspect of the invention relates to a first
generation (F1) hybrid soybean seed produced by crossing a plant of
the soybean variety A1016008 to a second soybean plant. Also included in
the invention are the F1 hybrid soybean plants grown from the hybrid
seed produced by crossing the soybean variety A1016008 to a second
soybean plant. Still further included in the invention are the seeds of
an F1 hybrid plant produced with the soybean variety A1016008 as one
parent, the second generation (F2) hybrid soybean plant grown from
the seed of the F1 hybrid plant, and the seeds of the F2 hybrid
plant.

[0011]Still yet another aspect of the invention is a method of producing
soybean seeds comprising crossing a plant of the soybean variety A1016008
to any second soybean plant, including itself or another plant of the
variety A1016008. In particular embodiments of the invention, the method
of crossing comprises the steps of a) planting seeds of the soybean
variety A1016008; b) cultivating soybean plants resulting from said seeds
until said plants bear flowers; c) allowing fertilization of the flowers
of said plants; and, d) harvesting seeds produced from said plants.

[0012]Still yet another aspect of the invention is a method of producing
hybrid soybean seeds comprising crossing the soybean variety A1016008 to
a second, distinct soybean plant which is nonisogenic to the soybean
variety A1016008. In particular embodiments of the invention, the
crossing comprises the steps of a) planting seeds of soybean variety
A1016008 and a second, distinct soybean plant, b) cultivating the soybean
plants grown from the seeds until the plants bear flowers; c) cross
pollinating a flower on one of the two plants with the pollen of the
other plant, and d) harvesting the seeds resulting from the cross
pollinating.

[0013]Still yet another aspect of the invention is a method for developing
a soybean plant in a soybean breeding program comprising: obtaining a
soybean plant, or its parts, of the variety A1016008; and b) employing
said plant or parts as a source of breeding material using plant breeding
techniques. In the method, the plant breeding techniques may be selected
from the group consisting of recurrent selection, mass selection, bulk
selection, backcrossing, pedigree breeding, genetic marker-assisted
selection and genetic transformation. In certain embodiments of the
invention, the soybean plant of variety A1016008 is used as the male or
female parent.

[0014]Still yet another aspect of the invention is a method of producing a
soybean plant derived from the soybean variety A1016008, the method
comprising the steps of: (a) preparing a progeny plant derived from
soybean variety A1016008 by crossing a plant of the soybean variety
A1016008 with a second soybean plant; and (b) crossing the progeny plant
with itself or a second plant to produce a progeny plant of a subsequent
generation which is derived from a plant of the soybean variety A1016008.
In one embodiment of the invention, the method further comprises: (c)
crossing the progeny plant of a subsequent generation with itself or a
second plant; and (d) repeating steps (b) and (c) for, for example, at
least 2, 3, 4 or more additional generations to produce an inbred soybean
plant derived from the soybean variety A1016008. Also provided by the
invention is a plant produced by this and the other methods of the
invention.

[0015]In another embodiment of the invention, the method of producing a
soybean plant derived from the soybean variety A1016008 further
comprises: (a) crossing the soybean variety A1016008-derived soybean
plant with itself or another soybean plant to yield additional soybean
variety A1016008-derived progeny soybean seed; (b) growing the progeny
soybean seed of step (a) under plant growth conditions, to yield
additional soybean variety A1016008-derived soybean plants; and (c)
repeating the crossing and growing steps of (a) and (b) to generate
further soybean variety A1016008-derived soybean plants. In specific
embodiments, steps (a) and (b) may be repeated at least 1, 2, 3, 4, or 5
or more times as desired. The invention still further provides a soybean
plant produced by this and the foregoing methods.

DETAILED DESCRIPTION OF THE INVENTION

[0016]The instant invention provides methods and composition relating to
plants, seeds and derivatives of the soybean variety A1016008. Soybean
variety A1016008 is adapted to mid to late group II growing regions.
Soybean variety A1016008 was developed from an initial cross between
CAK2903L0R*2 and GM_A19788. The breeding history of the variety can be
summarized as follows:

[0019]The soybean variety A1016008 has been judged to be uniform for
breeding purposes and testing. The variety A1016008 can be reproduced by
planting and growing seeds of the variety under self-pollinating or
sib-pollinating conditions, as is known to those of skill in the
agricultural arts. Variety A1016008 shows no variants other than what
would normally be expected due to environment or that would occur for
almost any characteristic during the course of repeated sexual
reproduction. The results of an objective evaluation of the variety are
presented below, in Table 1. Those of skill in the art will recognize
that these are typical values that may vary due to environment and that
other values that are substantially equivalent are within the scope of
the invention.

[0021]One aspect of the current invention concerns methods for crossing
the soybean variety A1016008 with itself or a second plant and the seeds
and plants produced by such methods. These methods can be used for
propagation of the soybean variety A1016008, or can be used to produce
hybrid soybean seeds and the plants grown therefrom. Hybrid soybean
plants can be used by farmers in the commercial production of soy
products or may be advanced in certain breeding protocols for the
production of novel soybean varieties. A hybrid plant can also be used as
a recurrent parent at any given stage in a backcrossing protocol during
the production of a single locus conversion of the soybean variety
A1016008.

[0022]Soybean variety A1016008 is well suited to the development of new
varieties based on the elite nature of the genetic background of the
variety. In selecting a second plant to cross with A1016008 for the
purpose of developing novel soybean varieties, it will typically be
desired to choose those plants which either themselves exhibit one or
more selected desirable characteristics or which exhibit the desired
characteristic(s) when in hybrid combination. Examples of potentially
desired characteristics include seed yield, lodging resistance,
emergence, seedling vigor, disease tolerance, maturity, plant height,
high oil content, high protein content and shattering resistance.

[0023]Choice of breeding or selection methods depends on the mode of plant
reproduction, the heritability of the trait(s) being improved, and the
type of variety used commercially (e.g., F1 hybrid variety, pureline
variety, etc.). For highly heritable traits, a choice of superior
individual plants evaluated at a single location will be effective,
whereas for traits with low heritability, selection should be based on
mean values obtained from replicated evaluations of families of related
plants. Popular selection methods commonly include pedigree selection,
modified pedigree selection, mass selection, recurrent selection and
backcrossing.

[0024]The complexity of inheritance influences choice of the breeding
method. Backcross breeding is used to transfer one or a few favorable
genes for a highly heritable trait into a desirable variety. This
approach has been used extensively for breeding disease-resistant
varieties (Bowers et al., 1992; Nickell and Bernard, 1992). Various
recurrent selection techniques are used to improve quantitatively
inherited traits controlled by numerous genes. The use of recurrent
selection in self-pollinating crops depends on the ease of pollination,
the frequency of successful hybrids from each pollination, and the number
of hybrid offspring from each successful cross.

[0025]Each breeding program should include a periodic, objective
evaluation of the efficiency of the breeding procedure. Evaluation
criteria vary depending on the goal and objectives, but should include
gain from selection per year based on comparisons to an appropriate
standard, overall value of the advanced breeding lines, and number of
successful varieties produced per unit of input (e.g., per year, per
dollar expended, etc.).

[0026]Promising advanced breeding lines are thoroughly tested and compared
to appropriate standards in environments representative of the commercial
target area(s) for generally three or more years. The best lines are
candidates for new commercial varieties. Those still deficient in a few
traits may be used as parents to produce new populations for further
selection.

[0027]These processes, which lead to the final step of marketing and
distribution, may take as much as eight to 12 years from the time the
first cross is made. Therefore, development of new varieties is a
time-consuming process that requires precise forward planning, efficient
use of resources, and a minimum of changes in direction.

[0028]A most difficult task is the identification of individuals that are
genetically superior, because for most traits the true genotypic value is
masked by other confounding plant traits or environmental factors. One
method of identifying a superior plant is to observe its performance
relative to other experimental plants and to one or more widely grown
standard varieties. Single observations are generally inconclusive, while
replicated observations provide a better estimate of genetic worth.

[0029]The goal of plant breeding is to develop new, unique and superior
soybean varieties and hybrids. The breeder initially selects and crosses
two or more parental lines, followed by repeated selfing and selection,
producing many new genetic combinations. Each year, the plant breeder
selects the germplasm to advance to the next generation. This germplasm
is grown under unique and different geographical, climatic and soil
conditions, and further selections are then made, during and at the end
of the growing season. The varieties which are developed are
unpredictable. This unpredictability is because the breeder's selection
occurs in unique environments, with no control at the DNA level (using
conventional breeding procedures), and with millions of different
possible genetic combinations being generated. A breeder of ordinary
skill in the art cannot predict the final resulting lines he develops,
except possibly in a very gross and general fashion. The same breeder
cannot produce the same variety twice by using the exact same original
parents and the same selection techniques. This unpredictability results
in the expenditure of large amounts of research monies to develop
superior new soybean varieties.

[0030]Pedigree breeding and recurrent selection breeding methods are used
to develop varieties from breeding populations. Breeding programs combine
desirable traits from two or more varieties or various broad-based
sources into breeding pools from which varieties are developed by selfing
and selection of desired phenotypes. The new varieties are evaluated to
determine which have commercial potential.

[0031]Pedigree breeding is commonly used for the improvement of
self-pollinating crops. Two parents which possess favorable,
complementary traits are crossed to produce an F1. An F2
population is produced by selfing one or several F1's. Selection of
the best individuals may begin in the F2 population (or later
depending upon the breeder's objectives); then, beginning in the F3,
the best individuals in the best families can be selected. Replicated
testing of families can begin in the F3 or F4 generation to
improve the effectiveness of selection for traits with low heritability.
At an advanced stage of inbreeding (i.e., F6 and F7), the best
lines or mixtures of phenotypically similar lines are tested for
potential release as new varieties.

[0032]Mass and recurrent selections can be used to improve populations of
either self- or cross-pollinating crops. A genetically variable
population of heterozygous individuals is either identified or created by
intercrossing several different parents. The best plants are selected
based on individual superiority, outstanding progeny, or excellent
combining ability. The selected plants are intercrossed to produce a new
population in which further cycles of selection are continued.

[0033]Backcross breeding has been used to transfer genetic loci for simply
inherited, highly heritable traits into a desirable homozygous variety
which is the recurrent parent. The source of the trait to be transferred
is called the donor or nonrecurrent parent. The resulting plant is
expected to have the attributes of the recurrent parent (i.e., variety)
and the desirable trait transferred from the donor parent. After the
initial cross, individuals possessing the phenotype of the donor parent
are selected and repeatedly crossed (backcrossed) to the recurrent
parent. The resulting plant is expected to have the attributes of the
recurrent parent (i.e., variety) and the desirable trait transferred from
the donor parent.

[0034]The single-seed descent procedure in the strict sense refers to
planting a segregating population, harvesting a sample of one seed per
plant, and using the one-seed sample to plant the next generation. When
the population has been advanced from the F2 to the desired level of
inbreeding, the plants from which lines are derived will each trace to
different F2 individuals. The number of plants in a population
declines each generation due to failure of some seeds to germinate or
some plants to produce at least one seed. As a result, not all of the
F2 plants originally sampled in the population will be represented
by a progeny when generation advance is completed.

[0035]In a multiple-seed procedure, soybean breeders commonly harvest one
or more pods from each plant in a population and thresh them together to
form a bulk. Part of the bulk is used to plant the next generation and
part is put in reserve. The procedure has been referred to as modified
single-seed descent or the pod-bulk technique.

[0036]The multiple-seed procedure has been used to save labor at harvest.
It is considerably faster to thresh pods with a machine than to remove
one seed from each by hand for the single-seed procedure. The
multiple-seed procedure also makes it possible to plant the same number
of seeds of a population each generation of inbreeding. Enough seeds are
harvested to make up for those plants that did not germinate or produce
seed.

[0037]Descriptions of other breeding methods that are commonly used for
different traits and crops can be found in one of several reference books
(e.g., Allard, 1960; Simmonds, 1979; Sneep et al., 1979; Fehr, 1987a,b).

[0038]Proper testing should detect any major faults and establish the
level of superiority or improvement over current varieties. In addition
to showing superior performance, there must be a demand for a new variety
that is compatible with industry standards or which creates a new market.
The introduction of a new variety will incur additional costs to the seed
producer, the grower, processor and consumer; for special advertising and
marketing, altered seed and commercial production practices, and new
product utilization. The testing preceding release of a new variety
should take into consideration research and development costs as well as
technical superiority of the final variety. For seed-propagated
varieties, it must be feasible to produce seed easily and economically.

[0039]Any time the soybean variety A1016008 is crossed with another,
different, variety, first generation (F1) soybean progeny are
produced. The hybrid progeny are produced regardless of characteristics
of the two varieties produced. As such, an F1 hybrid soybean plant
may be produced by crossing A1016008 with any second soybean plant. The
second soybean plant may be genetically homogeneous (e.g., inbred) or may
itself be a hybrid. Therefore, any F1 hybrid soybean plant produced
by crossing soybean variety A1016008 with a second soybean plant is a
part of the present invention.

[0040]Soybean plants (Glycine max L.) can be crossed by either natural or
mechanical techniques (see, e.g., Fehr, 1980). Natural pollination occurs
in soybeans either by self pollination or natural cross pollination,
which typically is aided by pollinating organisms. In either natural or
artificial crosses, flowering and flowering time are an important
consideration. Soybean is a short-day plant, but there is considerable
genetic variation for sensitivity to photoperiod (Hamner, 1969; Criswell
and Hume, 1972). The critical day length for flowering ranges from about
13 h for genotypes adapted to tropical latitudes to 24 h for
photoperiod-insensitive genotypes grown at higher latitudes (Shibles et
al., 1975). Soybeans seem to be insensitive to day length for 9 days
after emergence. Photoperiods shorter than the critical day length are
required for 7 to 26 days to complete flower induction (Borthwick and
Parker, 1938; Shanmugasundaram and Tsou, 1978).

[0041]Sensitivity to day length is an important consideration when
genotypes are grown outside of their area of adaptation. When genotypes
adapted to tropical latitudes are grown in the field at higher latitudes,
they may not mature before frost occurs. Plants can be induced to flower
and mature earlier by creating artificially short days or by grafting
(Fehr, 1980). Soybeans frequently are grown in winter nurseries located
at sea level in tropical latitudes where day lengths are much shorter
than their critical photoperiod. The short day lengths and warm
temperatures encourage early flowering and seed maturation, and genotypes
can produce a seed crop in 90 days or fewer after planting. Early
flowering is useful for generation advance when only a few
self-pollinated seeds per plant are needed, but not for artificial
hybridization because the flowers self-pollinate before they are large
enough to manipulate for hybridization. Artificial lighting can be used
to extend the natural day length to about 14.5 h to obtain flowers
suitable for hybridization and to increase yields of self-pollinated
seed.

[0042]The effect of a short photoperiod on flowering and seed yield can be
partly offset by altitude, probably due to the effects of cool
temperature (Major et al., 1975). At tropical latitudes, varieties
adapted to the northern U.S. perform more like those adapted to the
southern U.S. at high altitudes than they do at sea level.

[0043]The light level required to delay flowering is dependent on the
quality of light emitted from the source and the genotype being grown.
Blue light with a wavelength of about 480 nm requires more than 30 times
the energy to inhibit flowering as red light with a wavelength of about
640 nm (Parker et al., 1946).

[0044]Temperature can also play a significant role in the flowering and
development of soybean (Major et al., 1975). It can influence the time of
flowering and suitability of flowers for hybridization. Temperatures
below 21° C. or above 32° C. can reduce floral initiation
or seed set (Hamner, 1969; van Schaik and Probst, 1958). Artificial
hybridization is most successful between 26° C. and 32° C.
because cooler temperatures reduce pollen shed and result in flowers that
self-pollinate before they are large enough to manipulate. Warmer
temperatures frequently are associated with increased flower abortion
caused by moisture stress; however, successful crosses are possible at
about 35° C. if soil moisture is adequate.

[0045]Soybeans have been classified as indeterminate, semi-determinate,
and determinate based on the abruptness of stem termination after
flowering begins (Bernard and Weiss, 1973). When grown at their latitude
of adaptation, indeterminate genotypes flower when about one-half of the
nodes on the main stem have developed. They have short racemes with few
flowers, and their terminal node has only a few flowers. Semi-determinate
genotypes also flower when about one-half of the nodes on the main stem
have developed, but node development and flowering on the main stem stops
more abruptly than on indeterminates. Their racemes are short and have
few flowers, except for the terminal one, which may have several times
more flowers than those lower on the plant. Determinate varieties begin
flowering when all or most of the nodes on the main stem have developed.
They usually have elongated racemes that may be several centimeters in
length and may have a large number of flowers. Stem termination and
flowering habit are reported to be controlled by two major genes (Bernard
and Weiss, 1973).

[0046]Soybean flowers typically are self-pollinated on the day the corolla
opens. The amount of natural crossing, which is typically associated with
insect vectors such as honeybees, is approximately 1% for adjacent plants
within a row and 0.5% between plants in adjacent rows. The structure of
soybean flowers is similar to that of other legume species and consists
of a calyx with five sepals, a corolla with five petals, 10 stamens, and
a pistil (Carlson, 1973). The calyx encloses the corolla until the day
before anthesis. The corolla emerges and unfolds to expose a standard,
two wing petals, and two keel petals. An open flower is about 7 mm long
from the base of the calyx to the tip of the standard and 6 mm wide
across the standard. The pistil consists of a single ovary that contains
one to five ovules, a style that curves toward the standard, and a
club-shaped stigma. The stigma is receptive to pollen about 1 day before
anthesis and remains receptive for 2 days after anthesis, if the flower
petals are not removed. Filaments of nine stamens are fused, and the one
nearest the standard is free. The stamens form a ring below the stigma
until about 1 day before anthesis, then their filaments begin to elongate
rapidly and elevate the anthers around the stigma. The anthers dehisce on
the day of anthesis, pollen grains fall on the stigma, and within 10 h
the pollen tubes reach the ovary and fertilization is completed (Johnson
and Bernard, 1963).

[0047]Self-pollination occurs naturally in soybean with no manipulation of
the flowers. For the crossing of two soybean plants, it is typically
preferable, although not required, to utilize artificial hybridization.
In artificial hybridization, the flower used as a female in a cross is
manually cross pollinated prior to maturation of pollen from the flower,
thereby preventing self fertilization, or alternatively, the male parts
of the flower are emasculated using a technique known in the art.
Techniques for emasculating the male parts of a soybean flower include,
for example, physical removal of the male parts, use of a genetic factor
conferring male sterility, and application of a chemical gametocide to
the male parts.

[0048]For artificial hybridization employing emasculation, flowers that
are expected to open the following day are selected on the female parent.
The buds are swollen and the corolla is just visible through the calyx or
has begun to emerge. Usually no more than two buds on a parent plant are
prepared, and all self-pollinated flowers or immature buds are removed
with forceps. Special care is required to remove immature buds that are
hidden under the stipules at the leaf axil, and which could develop into
flowers at a later date. The flower is grasped between the thumb and
index finger and the location of the stigma determined by examining the
sepals. A long, curvy sepal covers the keel, and the stigma is on the
opposite side of the flower. The calyx is removed by grasping a sepal
with the forceps, pulling it down and around the flower, and repeating
the procedure until the five sepals are removed. The exposed corolla is
removed by grasping it just above the calyx scar, then lifting and
wiggling the forceps simultaneously. Care is taken to grasp the corolla
low enough to remove the keel petals without injuring the stigma. The
ring of anthers is visible after the corolla is removed, unless the
anthers were removed with the petals. Cross-pollination can then be
carried out using, for example, petri dishes or envelopes in which male
flowers have been collected. Desiccators containing calcium chloride
crystals are used in some environments to dry male flowers to obtain
adequate pollen shed.

[0049]It has been demonstrated that emasculation is unnecessary to prevent
self-pollination (Walker et al., 1979). When emasculation is not used,
the anthers near the stigma frequently are removed to make it clearly
visible for pollination. The female flower usually is hand-pollinated
immediately after it is prepared; although a delay of several hours does
not seem to reduce seed set. Pollen shed typically begins in the morning
and may end when temperatures are above 30° C., or may begin later
and continue throughout much of the day with more moderate temperatures.

[0050]Pollen is available from a flower with a recently opened corolla,
but the degree of corolla opening associated with pollen shed may vary
during the day. In many environments, it is possible to collect male
flowers and use them immediately without storage. In the southern U.S.
and other humid climates, pollen shed occurs in the morning when female
flowers are more immature and difficult to manipulate than in the
afternoon, and the flowers may be damp from heavy dew. In those
circumstances, male flowers are collected into envelopes or petri dishes
in the morning and the open container is typically placed in a desiccator
for about 4 h at a temperature of about 25° C. The desiccator may
be taken to the field in the afternoon and kept in the shade to prevent
excessive temperatures from developing within it. Pollen viability can be
maintained in flowers for up to 2 days when stored at about 5° C.
In a desiccator at 3° C., flowers can be stored successfully for
several weeks; however, varieties may differ in the percentage of pollen
that germinates after long-term storage (Kuehl, 1961).

[0051]Either with or without emasculation of the female flower, hand
pollination can be carried out by removing the stamens and pistil with a
forceps from a flower of the male parent and gently brushing the anthers
against the stigma of the female flower. Access to the stamens can be
achieved by removing the front sepal and keel petals, or piercing the
keel with closed forceps and allowing them to open to push the petals
away. Brushing the anthers on the stigma causes them to rupture, and the
highest percentage of successful crosses is obtained when pollen is
clearly visible on the stigma. Pollen shed can be checked by tapping the
anthers before brushing the stigma. Several male flowers may have to be
used to obtain suitable pollen shed when conditions are unfavorable, or
the same male may be used to pollinate several flowers with good pollen
shed.

[0052]When male flowers do not have to be collected and dried in a
desiccator, it may be desired to plant the parents of a cross adjacent to
each other. Plants usually are grown in rows 65 to 100 cm apart to
facilitate movement of personnel within the field nursery. Yield of
self-pollinated seed from an individual plant may range from a few seeds
to more than 1,000 as a function of plant density. A density of 30
plants/m of row can be used when 30 or fewer seeds per plant is adequate,
10 plants/m can be used to obtain about 100 seeds/plant, and 3 plants/m
usually results in maximum seed production per plant. Densities of 12
plants/m or less commonly are used for artificial hybridization.

[0053]Multiple planting dates about 7 to 14 days apart usually are used to
match parents of different flowering dates. When differences in flowering
dates are extreme between parents, flowering of the later parent can be
hastened by creating an artificially short day or flowering of the
earlier parent can be delayed by use of artificially long days or delayed
planting. For example, crosses with genotypes adapted to the southern
U.S. are made in northern U.S. locations by covering the late genotype
with a box, large can, or similar container to create an artificially
short photoperiod of about 12 h for about 15 days beginning when there
are three nodes with trifoliate leaves on the main stem. Plants induced
to flower early tend to have flowers that self-pollinate when they are
small and can be difficult to prepare for hybridization.

[0054]Grafting can be used to hasten the flowering of late flowering
genotypes. A scion from a late genotype grafted on a stock that has begun
to flower will begin to bloom up to 42 days earlier than normal (Kiihl et
al., 1977). First flowers on the scion appear from 21 to 50 days after
the graft.

[0055]Observing pod development 7 days after pollination generally is
adequate to identify a successful cross. Abortion of pods and seeds can
occur several weeks after pollination, but the percentage of abortion
usually is low if plant stress is minimized (Shibles et al., 1975). Pods
that develop from artificial hybridization can be distinguished from
self-pollinated pods by the presence of the calyx scar, caused by removal
of the sepals. The sepals begin to fall off as the pods mature;
therefore, harvest should be completed at or immediately before the time
the pods reach their mature color. Harvesting pods early also avoids any
loss by shattering.

[0056]Once harvested, pods are typically air-dried at not more than
38° C. until the seeds contain 13% moisture or less, then the
seeds are removed by hand. Seed can be stored satisfactorily at about
25° C. for up to a year if relative humidity is 50% or less. In
humid climates, germination percentage declines rapidly unless the seed
is dried to 7% moisture and stored in an air-tight container at room
temperature. Long-term storage in any climate is best accomplished by
drying seed to 7% moisture and storing it at 10° C. or less in a
room maintained at 50% relative humidity or in an air-tight container.

II. Further Embodiments of the Invention

[0057]In certain aspects of the invention, plants of soybean variety
A1016008 are provided modified to include at least a first desired
heritable trait. Such plants may, in one embodiment, be developed by a
plant breeding technique called backcrossing, wherein essentially all of
the morphological and physiological characteristics of a variety are
recovered in addition to a genetic locus transferred into the plant via
the backcrossing technique. By essentially all of the morphological and
physiological characteristics, it is meant that the characteristics of a
plant are recovered that are otherwise present when compared in the same
environment, other than an occasional variant trait that might arise
during backcrossing or direct introduction of a transgene. It is
understood that a locus introduced by backcrossing may or may not be
transgenic in origin, and thus the term backcrossing specifically
includes backcrossing to introduce loci that were created by genetic
transformation.

[0058]In a typical backcross protocol, the original variety of interest
(recurrent parent) is crossed to a second variety (nonrecurrent parent)
that carries the single locus of interest to be transferred. The
resulting progeny from this cross are then crossed again to the recurrent
parent and the process is repeated until a soybean plant is obtained
wherein essentially all of the desired morphological and physiological
characteristics of the recurrent parent are recovered in the converted
plant, in addition to the single transferred locus from the nonrecurrent
parent.

[0059]The selection of a suitable recurrent parent is an important step
for a successful backcrossing procedure. The goal of a backcross protocol
is to alter or substitute a single trait or characteristic in the
original variety. To accomplish this, a single locus of the recurrent
variety is modified or substituted with the desired locus from the
nonrecurrent parent, while retaining essentially all of the rest of the
desired genetic, and therefore the desired physiological and
morphological constitution of the original variety. The choice of the
particular nonrecurrent parent will depend on the purpose of the
backcross; one of the major purposes is to add some commercially
desirable, agronomically important trait to the plant. The exact
backcrossing protocol will depend on the characteristic or trait being
altered to determine an appropriate testing protocol. Although
backcrossing methods are simplified when the characteristic being
transferred is a dominant allele, a recessive allele may also be
transferred. In this instance it may be necessary to introduce a test of
the progeny to determine if the desired characteristic has been
successfully transferred.

[0060]Soybean varieties can also be developed from more than two parents
(Fehr, 1987a). The technique, known as modified backcrossing, uses
different recurrent parents during the backcrossing. Modified
backcrossing may be used to replace the original recurrent parent with a
variety having certain more desirable characteristics or multiple parents
may be used to obtain different desirable characteristics from each.

[0061]Many single locus traits have been identified that are not regularly
selected for in the development of a new inbred but that can be improved
by backcrossing techniques. Single locus traits may or may not be
transgenic; examples of these traits include, but are not limited to,
male sterility, herbicide resistance, resistance to bacterial, fungal, or
viral disease, insect resistance, restoration of male fertility, enhanced
nutritional quality, yield stability, and yield enhancement. These
comprise genes generally inherited through the nucleus.

[0062]Direct selection may be applied where the single locus acts as a
dominant trait. An example of a dominant trait is the herbicide
resistance trait. For this selection process, the progeny of the initial
cross are sprayed with the herbicide prior to the backcrossing. The
spraying eliminates any plants which do not have the desired herbicide
resistance characteristic, and only those plants which have the herbicide
resistance gene are used in the subsequent backcross. This process is
then repeated for all additional backcross generations.

[0063]Selection of soybean plants for breeding is not necessarily
dependent on the phenotype of a plant and instead can be based on genetic
investigations. For example, one may utilize a suitable genetic marker
which is closely genetically linked to a trait of interest. One of these
markers may therefore be used to identify the presence or absence of a
trait in the offspring of a particular cross, and hence may be used in
selection of progeny for continued breeding. This technique may commonly
be referred to as marker assisted selection. Any other type of genetic
marker or other assay which is able to identify the relative presence or
absence of a trait of interest in a plant may also be useful for breeding
purposes. Procedures for marker assisted selection applicable to the
breeding of soybeans are well known in the art. Such methods will be of
particular utility in the case of recessive traits and variable
phenotypes, or where conventional assays may be more expensive, time
consuming or otherwise disadvantageous. Types of genetic markers which
could be used in accordance with the invention include, but are not
necessarily limited to, Simple Sequence Length Polymorphisms (SSLPs)
(Williams et al., 1990), Randomly Amplified Polymorphic DNAs (RAPDs), DNA
Amplification Fingerprinting (DAF), Sequence Characterized Amplified
Regions (SCARs), Arbitrary Primed Polymerase Chain Reaction (AP-PCR),
Amplified Fragment Length Polymorphisms (AFLPs) (EP 534 858, specifically
incorporated herein by reference in its entirety), and Single Nucleotide
Polymorphisms (SNPs) (Wang et al., 1998).

[0064]Many qualitative characters also have potential use as
phenotype-based genetic markers in soybeans; however, some or many may
not differ among varieties commonly used as parents (Bernard and Weiss,
1973). The most widely used genetic markers are flower color (purple
dominant to white), pubescence color (brown dominant to gray), and pod
color (brown dominant to tan). The association of purple hypocotyl color
with purple flowers and green hypocotyl color with white flowers is
commonly used to identify hybrids in the seedling stage. Differences in
maturity, height, hilum color, and pest resistance between parents can
also be used to verify hybrid plants.

[0065]Many useful traits that can be introduced by backcrossing, as well
as directly intro a plant, are those which are introduced by genetic
transformation techniques. Genetic transformation may therefore be used
to insert a selected transgene into the soybean variety of the invention
or may, alternatively, be used for the preparation of transgenes which
can be introduced by backcrossing. Methods for the transformation of many
economically important plants, including soybeans, are well known to
those of skill in the art. Techniques which may be employed for the
genetic transformation of soybeans include, but are not limited to,
electroporation, microprojectile bombardment, Agrobacterium-mediated
transformation and direct DNA uptake by protoplasts.

[0066]To effect transformation by electroporation, one may employ either
friable tissues, such as a suspension culture of cells or embryogenic
callus or alternatively one may transform immature embryos or other
organized tissue directly. In this technique, one would partially degrade
the cell walls of the chosen cells by exposing them to pectin-degrading
enzymes (pectolyases) or mechanically wound tissues in a controlled
manner.

[0067]Protoplasts may also be employed for electroporation transformation
of plants (Bates, 1994; Lazzeri, 1995). For example, the generation of
transgenic soybean plants by electroporation of cotyledon-derived
protoplasts was described by Dhir and Widholm in Intl. Patent Appl. Publ.
No. WO 92/17598, the disclosure of which is specifically incorporated
herein by reference.

[0068]A particularly efficient method for delivering transforming DNA
segments to plant cells is microprojectile bombardment. In this method,
particles are coated with nucleic acids and delivered into cells by a
propelling force. Exemplary particles include those comprised of
tungsten, platinum, and preferably, gold. For the bombardment, cells in
suspension are concentrated on filters or solid culture medium.
Alternatively, immature embryos or other target cells may be arranged on
solid culture medium. The cells to be bombarded are positioned at an
appropriate distance below the macroprojectile stopping plate.

[0069]An illustrative embodiment of a method for delivering DNA into plant
cells by acceleration is the Biolistics Particle Delivery System, which
can be used to propel particles coated with DNA or cells through a
screen, such as a stainless steel or Nytex screen, onto a surface covered
with target soybean cells. The screen disperses the particles so that
they are not delivered to the recipient cells in large aggregates. It is
believed that a screen intervening between the projectile apparatus and
the cells to be bombarded reduces the size of the projectile aggregate
and may contribute to a higher frequency of transformation by reducing
the damage inflicted on the recipient cells by projectiles that are too
large.

[0070]Microprojectile bombardment techniques are widely applicable, and
may be used to transform virtually any plant species. The application of
microprojectile bombardment for the transformation of soybeans is
described, for example, in U.S. Pat. No. 5,322,783, the disclosure of
which is specifically incorporated herein by reference in its entirety.

[0071]Agrobacterium-mediated transfer is another widely applicable system
for introducing gene loci into plant cells. An advantage of the technique
is that DNA can be introduced into whole plant tissues, thereby bypassing
the need for regeneration of an intact plant from a protoplast. Modern
Agrobacterium transformation vectors are capable of replication in E.
coli as well as Agrobacterium, allowing for convenient manipulations
(Klee et al., 1985). Moreover, recent technological advances in vectors
for Agrobacterium-mediated gene transfer have improved the arrangement of
genes and restriction sites in the vectors to facilitate the construction
of vectors capable of expressing various polypeptide coding genes. The
vectors described have convenient multi-linker regions flanked by a
promoter and a polyadenylation site for direct expression of inserted
polypeptide coding genes. Additionally, Agrobacterium containing both
armed and disarmed Ti genes can be used for transformation.

[0072]In those plant strains where Agrobacterium-mediated transformation
is efficient, it is the method of choice because of the facile and
defined nature of the gene locus transfer. The use of
Agrobacterium-mediated plant integrating vectors to introduce DNA into
plant cells is well known in the art (Fraley et al., 1985; U.S. Pat. No.
5,563,055). Use of Agrobacterium in the context of soybean transformation
has been described, for example, by Chee and Slightom (1995) and in U.S.
Pat. No. 5,569,834, the disclosures of which are specifically
incorporated herein by reference in their entirety.

[0074]Many hundreds if not thousands of different genes are known and
could potentially be introduced into a soybean plant according to the
invention. Non-limiting examples of particular genes and corresponding
phenotypes one may choose to introduce into a soybean plant are presented
below.

[0075]A. Herbicide Resistance

[0076]Numerous herbicide resistance genes are known and may be employed
with the invention. An example is a gene conferring resistance to a
herbicide that inhibits the growing point or meristem, such as an
imidazalinone or a sulfonylurea. Exemplary genes in this category code
for mutant ALS and AHAS enzyme as described, for example, by Lee et al.,
(1988); Gleen et al., (1992) and Miki et al., (1990).

[0077]Resistance genes for glyphosate (resistance conferred by mutant
5-enolpyruvl-3 phosphikimate synthase (EPSPS) and aroA genes,
respectively) and other phosphono compounds such as glufosinate
(phosphinothricin acetyl transferase (PAT) and Streptomyces hygroscopicus
phosphinothricin-acetyl transferase (bar) genes) may also be used. See,
for example, U.S. Pat. No. 4,940,835 to Shah, et al., which discloses the
nucleotide sequence of a form of EPSPS which can confer glyphosate
resistance. Examples of specific EPSPS transformation events conferring
glyphosate resistance are provided by U.S. Pat. No. 6,040,497.

[0078]A DNA molecule encoding a mutant aroA gene can be obtained under
ATCC accession number 39256, and the nucleotide sequence of the mutant
gene is disclosed in U.S. Pat. No. 4,769,061 to Comai. European patent
application No. 0 333 033 to Kumada et al., and U.S. Pat. No. 4,975,374
to Goodman et al., disclose nucleotide sequences of glutamine synthetase
genes which confer resistance to herbicides such as L-phosphinothricin.
The nucleotide sequence of a phosphinothricin-acetyltransferase gene is
provided in European application No. 0 242 246 to Leemans et al. DeGreef
et al., (1989), describe the production of transgenic plants that express
chimeric bar genes coding for phosphinothricin acetyl transferase
activity. Exemplary of genes conferring resistance to phenoxy propionic
acids and cyclohexones, such as sethoxydim and haloxyfop are the Acct-S1,
Acct-S2 and Acct-S3 genes described by Marshall et al., (1992).

[0079]Genes are also known conferring resistance to a herbicide that
inhibits photosynthesis, such as a triazine (psbA and gs+ genes) and a
benzonitrile (nitrilase gene). Przibila et al., (1991), describe the
transformation of Chlamydomonas with plasmids encoding mutant psbA genes.
Nucleotide sequences for nitrilase genes are disclosed in U.S. Pat. No.
4,810,648 to Stalker, and DNA molecules containing these genes are
available under ATCC Accession Nos. 53435, 67441, and 67442. Cloning and
expression of DNA coding for a glutathione S-transferase is described by
Hayes et al., (1992).

[0080]US Patent Application No: 20030135879 describes isolation of a gene
for dicamba monooxygenase (DMO) from Psueodmonas maltophilia which is
involved in the conversion of a herbicidal form of the herbicide dicamba
to a non-toxic 3,6-dichlorosalicylic acid and thus may be used for
producing plants tolerant to this herbicide.

[0081]B. Disease Resistance

[0082]Plant defenses are often activated by specific interaction between
the product of a disease resistance gene (R) in the plant and the product
of a corresponding avirulence (Avr) gene in the pathogen. A plant line
can be transformed with cloned resistance gene to engineer plants that
are resistant to specific pathogen strains. See, for example Jones et
al., (1994) (cloning of the tomato Cf-9 gene for resistance to
Cladosporium fulvum); Martin et al., (1993) (tomato Pto gene for
resistance to Pseudomonas syringae pv. tomato); and Mindrinos et al.,
(1994) (Arabidopsis RPS2 gene for resistance to Pseudomonas syringae).

[0083]A viral-invasive protein or a complex toxin derived therefrom may
also be used for viral disease resistance. For example, the accumulation
of viral coat proteins in transformed plant cells imparts resistance to
viral infection and/or disease development effected by the virus from
which the coat protein gene is derived, as well as by related viruses.
See Beachy et al. (1990). Coat protein-mediated resistance has been
conferred upon transformed plants against alfalfa mosaic virus, cucumber
mosaic virus, tobacco streak virus, potato virus X, potato virus Y,
tobacco etch virus, tobacco rattle virus and tobacco mosaic virus. Id.

[0084]A virus-specific antibody may also be used. See, for example,
Tavladoraki et al. (1993), who show that transgenic plants expressing
recombinant antibody genes are protected from virus attack.

[0087]One example of an insect resistance gene includes a Bacillus
thuringiensis protein, a derivative thereof or a synthetic polypeptide
modeled thereon. See, for example, Geiser et al. (1986), who disclose the
cloning and nucleotide sequence of a Bacillus thuringiensis
δ-endotoxin gene. Moreover, DNA molecules encoding
δ-endotoxin genes can be purchased from the American Type Culture
Collection, Manassas, Va., for example, under ATCC Accession Nos. 40098,
67136, 31995 and 31998. Another example is a lectin. See, for example,
Van Damme et al., (1994), who disclose the nucleotide sequences of
several Clivia miniata mannose-binding lectin genes. A vitamin-binding
protein may also be used, such as avidin. See PCT application US93/06487,
the contents of which are hereby incorporated by reference. This
application teaches the use of avidin and avidin homologues as larvicides
against insect pests.

[0089]Still other examples include an insect-specific antibody or an
immunotoxin derived therefrom and a developmental-arrestive protein. See
Taylor et al., (1994), who described enzymatic inactivation in transgenic
tobacco via production of single-chain antibody fragments.

[0090]D. Male Sterility

[0091]Genetic male sterility is available in soybeans and can increase the
efficiency with which hybrids are made, in that it can eliminate the need
to physically emasculate the soybean plant used as a female in a given
cross. (Brim and Stuber, 1973). Herbicide-inducible male sterility
systems have also been described. (U.S. Pat. No. 6,762,344).

[0092]Where one desires to employ male-sterility systems, it may be
beneficial to also utilize one or more male-fertility restorer genes. For
example, where cytoplasmic male sterility (CMS) is used, hybrid seed
production requires three inbred lines: (1) a cytoplasmically
male-sterile line having a CMS cytoplasm; (2) a fertile inbred with
normal cytoplasm, which is isogenic with the CMS line for nuclear genes
("maintainer line"); and (3) a distinct, fertile inbred with normal
cytoplasm, carrying a fertility restoring gene ("restorer" line). The CMS
line is propagated by pollination with the maintainer line, with all of
the progeny being male sterile, as the CMS cytoplasm is derived from the
female parent. These male sterile plants can then be efficiently employed
as the female parent in hybrid crosses with the restorer line, without
the need for physical emasculation of the male reproductive parts of the
female parent.

[0093]The presence of a male-fertility restorer gene results in the
production of fully fertile F1 hybrid progeny. If no restorer gene
is present in the male parent, male-sterile hybrids are obtained. Such
hybrids are useful where the vegetative tissue of the soybean plant is
utilized, but in many cases the seeds will be deemed the most valuable
portion of the crop, so fertility of the hybrids in these crops must be
restored. Therefore, one aspect of the current invention concerns plants
of the soybean variety A1016008 comprising a genetic locus capable of
restoring male fertility in an otherwise male-sterile plant. Examples of
male-sterility genes and corresponding restorers which could be employed
with the plants of the invention are well known to those of skill in the
art of plant breeding (see, e.g., U.S. Pat. No. 5,530,191 and U.S. Pat.
No. 5,684,242, the disclosures of which are each specifically
incorporated herein by reference in their entirety).

[0096]Phytate metabolism may also be modified by introduction of a
phytase-encoding gene to enhance breakdown of phytate, adding more free
phosphate to the transformed plant. For example, see Van Hartingsveldt et
al., (1993), for a disclosure of the nucleotide sequence of an
Aspergillus niger phytase gene. In soybean, this, for example, could be
accomplished by cloning and then reintroducing DNA associated with the
single allele which is responsible for soybean mutants characterized by
low levels of phytic acid. See Raboy et al., (2000).

[0097]A number of genes are known that may be used to alter carbohydrate
metabolism. For example, plants may be transformed with a gene coding for
an enzyme that alters the branching pattern of starch. See Shiroza et
al., (1988) (nucleotide sequence of Streptococcus mutans
fructosyltransferase gene), Steinmetz et al., (1985) (nucleotide sequence
of Bacillus subtilis levansucrase gene), Pen et al., (1992) (production
of transgenic plants that express Bacillus licheniformis
α-amylase), Elliot et al., (1993) (nucleotide sequences of tomato
invertase genes), Sergaard et al., (1993) (site-directed mutagenesis of
barley α-amylase gene), and Fisher et al., (1993) (maize endosperm
starch branching enzyme II). The Z10 gene encoding a 10 kD zein storage
protein from maize may also be used to alter the quantities of 10 kD zein
in the cells relative to other components (Kirihara et al., 1988).

III. Origin and Breeding History of an Exemplary Single Locus Converted
Plant

[0098]It is known to those of skill in the art that, by way of the
technique of backcrossing, one or more traits may be introduced into a
given variety while otherwise retaining essentially all of the traits of
that variety. An example of such backcrossing to introduce a trait into a
starting variety is described in U.S. Pat. No. 6,140,556, the entire
disclosure of which is specifically incorporated herein by reference. The
procedure described in U.S. Pat. No. 6,140,556 can be summarized as
follows: The soybean variety known as Williams '82 [Glycine max L. Merr.]
(Reg. No. 222, PI 518671) was developed using backcrossing techniques to
transfer a locus comprising the Rps1 gene to the variety Williams
(Bernard and Cremeens, 1988). Williams '82 is a composite of four
resistant lines from the BC6F3 generation, which were selected
from 12 field-tested resistant lines from Williams×Kingwa. The
variety Williams was used as the recurrent parent in the backcross and
the variety Kingwa was used as the source of the Rps1 locus. This
gene locus confers resistance to 19 of the 24 races of the fungal agent
phytophthora rot.

[0099]The F1 or F2 seedlings from each backcross round were
tested for resistance to the fungus by hypocotyl inoculation using the
inoculum of race 5. The final generation was tested using inoculum of
races 1 to 9. In a backcross such as this, where the desired
characteristic being transferred to the recurrent parent is controlled by
a major gene which can be readily evaluated during the backcrossing, it
is common to conduct enough backcrosses to avoid testing individual
progeny for specific traits such as yield in extensive replicated tests.
In general, four or more backcrosses are used when there is no evaluation
of the progeny for specific traits, such as yield. As in this example,
lines with the phenotype of the recurrent parent may be composited
without the usual replicated tests for traits such as yield, protein or
oil percentage in the individual lines.

[0100]The variety Williams '82 is comparable to the recurrent parent
variety Williams in its traits except resistance to phytophthora rot. For
example, both varieties have a relative maturity of 38, indeterminate
stems, white flowers, brown pubescence, tan pods at maturity and shiny
yellow seeds with black to light black hila.

IV. Tissue Cultures and In Vitro Regeneration of Soybean Plants

[0101]A further aspect of the invention relates to tissue cultures of the
soybean variety designated A1016008. As used herein, the term "tissue
culture" indicates a composition comprising isolated cells of the same or
a different type or a collection of such cells organized into parts of a
plant. Exemplary types of tissue cultures are protoplasts, calli and
plant cells that are intact in plants or parts of plants, such as
embryos, pollen, flowers, leaves, roots, root tips, anthers, and the
like. In a preferred embodiment, the tissue culture comprises embryos,
protoplasts, meristematic cells, pollen, leaves or anthers.

[0102]Exemplary procedures for preparing tissue cultures of regenerable
soybean cells and regenerating soybean plants therefrom, are disclosed in
U.S. Pat. No. 4,992,375; U.S. Pat. No. 5,015,580; U.S. Pat. No.
5,024,944, and U.S. Pat. No. 5,416,011, each of the disclosures of which
is specifically incorporated herein by reference in its entirety.

[0103]An important ability of a tissue culture is the capability to
regenerate fertile plants. This allows, for example, transformation of
the tissue culture cells followed by regeneration of transgenic plants.
For transformation to be efficient and successful, DNA must be introduced
into cells that give rise to plants or germ-line tissue.

[0104]Soybeans typically are regenerated via two distinct processes: shoot
morphogenesis and somatic embryogenesis (Finer, 1996). Shoot
morphogenesis is the process of shoot meristem organization and
development. Shoots grow out from a source tissue and are excised and
rooted to obtain an intact plant. During somatic embryogenesis, an embryo
(similar to the zygotic embryo), containing both shoot and root axes, is
formed from somatic plant tissue. An intact plant rather than a rooted
shoot results from the germination of the somatic embryo.

[0105]Shoot morphogenesis and somatic embryogenesis are different
processes and the specific route of regeneration is primarily dependent
on the explant source and media used for tissue culture manipulations.
While the systems are different, both systems show variety-specific
responses where some lines are more responsive to tissue culture
manipulations than others. A line that is highly responsive in shoot
morphogenesis may not generate many somatic embryos. Lines that produce
large numbers of embryos during an `induction` step may not give rise to
rapidly-growing proliferative cultures. Therefore, it may be desired to
optimize tissue culture conditions for each soybean line. These
optimizations may readily be carried out by one of skill in the art of
tissue culture through small-scale culture studies. In addition to
line-specific responses, proliferative cultures can be observed with both
shoot morphogenesis and somatic embryogenesis. Proliferation is
beneficial for both systems, as it allows a single, transformed cell to
multiply to the point that it will contribute to germ-line tissue.

[0106]Shoot morphogenesis was first reported by Wright et al. (1986) as a
system whereby shoots were obtained de novo from cotyledonary nodes of
soybean seedlings. The shoot meristems were formed subepidermally and
morphogenic tissue could proliferate on a medium containing benzyl
adenine (BA). This system can be used for transformation if the
subepidermal, multicellular origin of the shoots is recognized and
proliferative cultures are utilized. The idea is to target tissue that
will give rise to new shoots and proliferate those cells within the
meristematic tissue to lessen problems associated with chimerism.
Formation of chimeras, resulting from transformation of only a single
cell in a meristem, are problematic if the transformed cell is not
adequately proliferated and does not does not give rise to germ-line
tissue. Once the system is well understood and reproduced satisfactorily,
it can be used as one target tissue for soybean transformation.

[0107]Somatic embryogenesis in soybean was first reported by Christianson
et al. (1983) as a system in which embryogenic tissue was initially
obtained from the zygotic embryo axis. These embryogenic cultures were
proliferative but the repeatability of the system was low and the origin
of the embryos was not reported. Later histological studies of a
different proliferative embryogenic soybean culture showed that
proliferative embryos were of apical or surface origin with a small
number of cells contributing to embryo formation. The origin of primary
embryos (the first embryos derived from the initial explant) is dependent
on the explant tissue and the auxin levels in the induction medium
(Hartweck et al., 1988). With proliferative embryonic cultures, single
cells or small groups of surface cells of the `older` somatic embryos
form the `newer` embryos.

[0108]Embryogenic cultures can also be used successfully for regeneration,
including regeneration of transgenic plants, if the origin of the embryos
is recognized and the biological limitations of proliferative embryogenic
cultures are understood. Biological limitations include the difficulty in
developing proliferative embryogenic cultures and reduced fertility
problems (culture-induced variation) associated with plants regenerated
from long-term proliferative embryogenic cultures. Some of these problems
are accentuated in prolonged cultures. The use of more recently cultured
cells may decrease or eliminate such problems.

V. Definitions

[0109]In the description and tables, a number of terms are used. In order
to provide a clear and consistent understanding of the specification and
claims, the following definitions are provided:

[0110]A: When used in conjunction with the word "comprising" or other open
language in the claims, the words "a" and "an" denote "one or more."

[0111]Allele: Any of one or more alternative forms of a gene locus, all of
which alleles relate to one trait or characteristic. In a diploid cell or
organism, the two alleles of a given gene occupy corresponding loci on a
pair of homologous chromosomes.

[0112]Backcrossing: A process in which a breeder repeatedly crosses hybrid
progeny, for example a first generation hybrid (F1), back to one of
the parents of the hybrid progeny. Backcrossing can be used to introduce
one or more single locus conversions from one genetic background into
another.

[0113]Brown Stem Rot: This is a visual disease score from 1 to 9 comparing
all genotypes in a given test. The score is based on leaf symptoms of
yellowing and necrosis caused by brown stem rot. A score of 1 indicates
no symptoms. Visual scores range to a score of 9 which indicates severe
symptoms of leaf yellowing and necrosis.

[0114]Chromatography: A technique wherein a mixture of dissolved
substances are bound to a solid support followed by passing a column of
fluid across the solid support and varying the composition of the fluid.
The components of the mixture are separated by selective elution.

[0115]Crossing: The mating of two parent plants.

[0116]Cross-pollination: Fertilization by the union of two gametes from
different plants.

[0117]Emasculate: The removal of plant male sex organs or the inactivation
of the organs with a cytoplasmic or nuclear genetic factor or a chemical
agent conferring male sterility.

[0118]Emergence: This is a score indicating the ability of a seed to
emerge from the soil after planting. Each genotype is given a 1 to 9
score based on its percent of emergence. A score of 1 indicates an
excellent rate and percent of emergence, an intermediate score of 5
indicates average ratings and a 9 score indicates a very poor rate and
percent of emergence.

[0119]Enzymes: Molecules which can act as catalysts in biological
reactions.

[0120]F1 Hybrid: The first generation progeny of the cross of two
nonisogenic plants.

[0121]Genotype: The genetic constitution of a cell or organism.

[0122]Haploid: A cell or organism having one set of the two sets of
chromosomes in a diploid.

[0123]Iron-Deficiency Chlorosis: A plant scoring system ranging from 1 to
9 based on visual observations. A score of 1 means no stunting of the
plants or yellowing of the leaves and a score of 9 indicates the plants
are dead or dying caused by iron-deficiency chlorosis; a score of 5 means
plants have intermediate health with some leaf yellowing.

[0124]Linkage: A phenomenon wherein alleles on the same chromosome tend to
segregate together more often than expected by chance if their
transmission was independent.

[0125]Lodging Resistance: Lodging is rated on a scale of 1 to 9. A score
of 1 indicates erect plants. A score of 5 indicates plants are leaning at
a 45 degree(s) angle in relation to the ground and a score of 9 indicates
plants are laying on the ground.

[0126]Marker: A readily detectable phenotype, preferably inherited in
codominant fashion (both alleles at a locus in a diploid heterozygote are
readily detectable), with no environmental variance component, i.e.,
heritability of 1.

[0127]Maturity Date: Plants are considered mature when 95% of the pods
have reached their mature color. The maturity date is typically described
in measured days after August 31 in the northern hemisphere.

[0128]Phenotype: The detectable characteristics of a cell or organism,
which characteristics are the manifestation of gene expression.

[0129]Phytophthora Tolerance: Tolerance to Phytophthora root rot is rated
on a scale of 1 to 9, with a score of 1 being the best or highest
tolerance ranging down to a score of 9, which indicates the plants have
no tolerance to Phytophthora.

[0130]Plant Height: Plant height is taken from the top of soil to the top
node of the plant and is measured in inches.

[0131]Regeneration: The development of a plant from tissue culture.

[0132]Relative Maturity: The maturity grouping designated by the soybean
industry over a given growing area. This figure is generally divided into
tenths of a relative maturity group. Within narrow comparisons, the
difference of a tenth of a relative maturity group equates very roughly
to a day difference in maturity at harvest.

[0133]Seed Protein Peroxidase Activity. Seed protein peroxidase activity
is defined as a chemical taxonomic technique to separate varieties based
on the presence or absence of the peroxidase enzyme in the seed coat.
There are two types of soybean varieties, those having high peroxidase
activity (dark red color) and those having low peroxidase activity (no
color).

[0134]Seed Yield (Bushels/Acre): The yield in bushels/acre is the actual
yield of the grain at harvest.

[0135]Self-pollination: The transfer of pollen from the anther to the
stigma of the same plant.

[0136]Shattering: The amount of pod dehiscence prior to harvest. Pod
dehiscence involves seeds falling from the pods to the soil. This is a
visual score from 1 to 9 comparing all genotypes within a given test. A
score of 1 means pods have not opened and no seeds have fallen out. A
score of 5 indicates approximately 50% of the pods have opened, with
seeds falling to the ground and a score of 9 indicates 100% of the pods
are opened.

[0137]Single Locus Converted (Conversion) Plant: Plants which are
developed by a plant breeding technique called backcrossing, wherein
essentially all of the morphological and physiological characteristics of
a soybean variety are recovered in addition to the characteristics of the
single locus transferred into the variety via the backcrossing technique
and/or by genetic transformation.

[0138]Substantially Equivalent: A characteristic that, when compared, does
not show a statistically significant difference (e.g., p=0.05) from the
mean.

[0139]Tissue Culture: A composition comprising isolated cells of the same
or a different type or a collection of such cells organized into parts of
a plant.

[0140]Transgene: A genetic locus comprising a sequence which has been
introduced into the genome of a soybean plant by transformation.

VI. Deposit Information

[0141]A deposit of the soybean variety A1016008, which is disclosed herein
above and referenced in the claims, will be made with the American Type
Culture Collection (ATCC), 10801 University Blvd., Manassas, Va.
20110-2209. The date of deposit is ______ and the accession number for
those deposited seeds of soybean variety A1016008 is ATCC Accession No.
______. All restrictions upon the deposit have been removed, and the
deposit is intended to meet all of the requirements of 37 C.F.R.
§1.801-1.809. The deposit will be maintained in the depository for a
period of 30 years, or 5 years after the last request, or for the
effective life of the patent, whichever is longer, and will be replaced
if necessary during that period.